No-carrier-added labeling of the neuroprotective Ebselen with selenium-73 and selenium-75

Article abstract of DOI:10.1002/jlcr.3274

Selenium-73 is a positron emitting non-standard radionuclide, which is suitable for positron emission tomography. A copper-catalyzed reaction allowed no-carrier-added labeling of the anti-inflammatory seleno-organic compound Ebselen with ⁷³Se and ⁷⁵Se under addition of sulfur carrier in a one-step reaction. The new authentically labeled radioselenium molecule is thus available for preclinical evaluation and positron emission tomography studies.

Full text of DOI:10.1002/jlcr.3274

Special Issue Article
Received 23 September 2014,
Revised 19 January 2015,
Accepted 20 January 2015
Published online in Wiley Online Library
(wileyonlinelibrary.com) DOI: 10.1002/jlcr.3274
No-carrier-added labeling of the neuropro-
tective Ebselen with selenium-73 and
selenium-75†,‡
*
Andreas Helfer, Johannes Ermert, Sven Humpert, and Heinz H. Coenen
Selenium-73 is a positron emitting non-standard radionuclide, which is suitable for positron emission tomography. A
copper-catalyzed reaction allowed no-carrier-added labeling of the anti-inflammatory seleno-organic compound Ebselen
with 73Se and 75Se under addition of sulfur carrier in a one-step reaction. The new authentically labeled radioselenium
molecule is thus available for preclinical evaluation and positron emission tomography studies.
Keywords: radioselenium; n.c.a. labeling; Ebselen; positron emission tomography
So far, most 75Se-labeled compounds were synthesized by
carrier-added (c.a.) methods. In consideration of the possible
Introduction
toxicity of selenium compounds, however, some labeling methods
for the preparation of radioseleno compounds have also been
developed at the n.c.a. level during the last three decades. The
following methods were described, which allow the synthesis of
n.c.a. radioselenium compounds: Starting from n.c.a. methyl[75Se]
selenide, which is available using sulfur as non-isotopic carrier,4
alkylation of disubstituted radio selenoureas11 and conversion of
alkyl[75Se]selenocyanates with alkyl and aryl lithium or Grignard
compounds were realized.12 Only the latter method allows the
synthesis of aromatic radioselenoethers.
Ebselen, 2-phenyl-1,2-benzisoselenazol-3(2H)-one (Figure 1), is
an organoselenium compound with glutathione peroxidase-like,
thiol-dependent, hydroperoxide reducing activity and
neuroprotective and cytoprotective properties.13,14 It inhibits
radically induced apoptosis while exhibiting low toxicity.
Furthermore, the aromatically bound selenium is highly stable
with respect to degradation in vivo. Ebselen was investigated
as a therapeutic agent for injury and stroke.15–17 Quite recently,
it was found that Ebselen also inhibits inositol monophosphatase
and induces lithium-like effects on mouse behavior.
Even being highly toxic, selenium is known as an essential trace
element for mammals.1 Its physiological role in mammals could
not have been elucidated without the use of radioselenium.2
Because of the lack of suitable isotopes of sulfur for in vivo
molecular imaging and the chemical homology of sulfur and
selenium, the positron emitter selenium-73 (t1/2 = 7.1h) may serve
as a possible substitute for sulfur to obtain analogous radiotracers
for in vivo application. A possible disadvantage of selenium-73 is its
radioactive, rather long-lived decay product 73As (t1/2 = 80days). Its
contribution to the radiation dose must of course be carefully
determined. This radiation dose depends on the individual kinetics
of the 73Se-tracer applied, as well as on those of the decay-
generated chemical forms of arsenic at the no-carrier-added
(n.c.a.) level, which has a strong influence on the excretion.3
However, no study on the speciation of radioarsenic formed
by the decay of 73Se in vivo has been performed so far.
Development of synthesis methods for 73Se-labeled radio tracers
is for convenience generally carried out with the long-lived
radioisotope selenium-75.4,5 Earlier, selenium-75 was even used as a
gamma ray emitting nuclide in a few radiopharmaceuticals6 despite
its decay characteristics that are in principle unsuitable for its in vivo
medical application.7 Nowadays, the use of selenium-75 is almost
restricted to biochemical in vitro studies.8 Its half-life of 120 days
allows to perform multi-step reactions with a single production
batch. 75Se can easily be produced via the 75As(p,n)75Se nuclear
reaction on small cyclotrons with a proton energy of below 18 MeV.
With the growing importance of positron emission tomography
(PET) for in vivo imaging, there is great interest to expand the list of
usable positron emitters such as selenium-73.9 Its nuclear
properties with Eβ+ = 1.3MeV and a β+ branching of 65% make this
isotope attractive for PET application, and the half-life of 7.1h
allows for the implementation of extended syntheses and imaging
protocols. However, the production via the 75As(p,3n)73Se nuclear
reaction necessitates proton energies of 30 to 40 MeV.10 The
nuclear reactions used for generation of both selenium isotopes
allow their production in n.c.a. form.
Institut für Neurowissenschaften und Medizin, INM-5: Nuklearchemie,
Forschungszentrum Jülich, 52425 Jülich, Germany
*Correspondence to: Heinz H. Coenen, Institut für Neurowissenschaften und
Medizin, INM-5: Nuklearchemie, Forschungszentrum Jülich, Jülich, Germany.
E-mail: h.h.coenen@fz-juelich.de
†This article is published in Journal of Labelled Compounds and Radiopharmaceuticals
as a special issue on ‘Bengt Långström’, edited by Antony Gee, Department of
Chemistry and Biology, Division of Imaging Sciences and Bioengineering, Kings College
London, UK and Albert Windhorst, Department of Radiology and Nuclear Medicine,
VU University Medical Center, Amsterdam, The Netherlands.
‡Dedicated to Prof. Dr. Bengt Långström, with deepest appreciation, to celebrate
his outstanding life-long contribution to the field of radiochemistry and on the
occasion of his retirement as editor of the Journal of Labelled Compounds and
Radiopharmaceuticals.
J. Label Compd. Radiopharm 2015, 58 141–145
Copyright © 2015 John Wiley & Sons, Ltd.

A. Helfer et al.
(Scheme 2).20 A new concept for synthesizing Ebselen on a
macroscopic scale uses a copper-catalyzed one-step reaction
resulting in good yields (cf. Scheme 4).21
Biography
Heinz H. Coenen received his PhD in
nuclear chemistry at the University of
Cologne in 1979 and began a career
in radiopharmaceutical chemistry with
a PostDoc position at the Research
Centre Jülich (FZJ). Since 1996, he is a
full professor at the University of
Cologne and a director at the Institute
of Nuclear Chemistry at FZJ. His
research interests focus especially on
the development of radionuclides and
radiotracers for in vivo molecular
imaging and their translation into clinical application. Besides
other duties, he acted as President of the Society of
Radiopharmaceutical Sciences.
N.c.a. [75Se]Ebselen via ortho-lithiation of
N-phenylbenzamide
The cyclotron-production of radioselenium isotopes by
irradiation of a Cu3As target yielded n.c.a. elemental selenium
as solution in benzene after thermochromatographic isolation
as described in detail before.10,11,22,23 [Attention must be paid
if the volume of this solution is reduced as it will lead to a loss
of reactive selenium]. The preparation of Ebselen as reference
material was performed as originally,20 using two equivalents
of n-butyllithium in tetrahydrofuran, followed by the addition
of elemental selenium powder at 0 °C each, and oxidative ring
closure with CuBr2 at À78 °C (Scheme 2). The radiosynthesis
according to Scheme 1 was attempted using a mixture of THF
and the selenium containing benzene. This solvent mixture
had a melting point just below 0 °C, and thus, it was much
higher than the temperature (À78 °C) of the oxidative ring
closure described in the original method. To perform the n.c.a.
radiosynthesis, N-phenylbenzamide was therefore first lithiated
in a separate vial using THF at 0 °C and then transferred to the
benzene solution containing the radioselenium. The subsequent
oxidation step with CuBr2 was also carried out at the same
reaction temperature. The organic product was separated from
water-soluble products by solid phase extraction and was eluted
with methanol. However, using these conditions, no formation of
n.c.a. [75Se]Ebselen was observed. Neither the addition of sulfur
as non-isotopic carrier nor the addition of selenium as carrier
gave the desired product.
Therefore, Ebselen can be considered as possible lithium
mimetic for the treatment of bipolar disorder.18 An earlier
authentic labeling of the molecule with selenium-75 was
published, however, in c.a. form only.19 In order to avoid
pharmacodynamic effects, new labeling methods had to be
developed for the production of the n.c.a. 73Se-labeled
radiopharmaceuticals.
Results and discussion
For convenience, the development and optimization of all
radiosyntheses described here were performed using the longer-
lived isotope selenium-75. The optimum reaction conditions found
were then adapted to the corresponding radiosynthesis using the
short-lived positron emitter selenium-73.
[
75Se]Ebselen via copper-catalyzed one-pot radiosynthesis
Synthesis of Ebselen and derivatives thereof was conventionally
performed via three routes. The bis(ortho-benzoic acid) diselenide According to the recently published synthetic method,21 the
route is a method in which anthranilic acid is converted into a transition metal-catalyzed coupling of selenium with 2-iodo-N-
heterocycle by a series of reactions. The first step is the conversion phenylbenzamide was performed with 20 mol % of CuI and
into the diselenide derivative using Na2Se2. This method was also 1,10-phenanthroline and selenium powder in the presence of
used for the synthesis of c.a. [75Se]Ebselen (Scheme 1).19
K2CO3 in DMF at 110 °C within 8 h. For work up, the reaction
However, it cannot be employed for the n.c.a. synthesis of mixture was treated with an aqueous solution.
[73Se]Ebselen and [75Se]Ebselen because of the necessity to form
The proposed mechanism of the Cu-catalyzed reaction is
a diselenide as intermediate that is not achievable without given in Scheme 3. A base-promoted reaction forms a Cu-amide
addition of carrier. The second method is based on lithiation of complex followed by insertion of selenium into the Cu–N bond,
benzanilide with n-butyllithium, subsequent reaction with and finally, reductive elimination of CuL leads to cyclization by
selenium powder, and finally, oxidative cyclization by copper forming the Se–C bond.21
The first attempts to synthesize radiolabeled Ebselen by this
procedure were performed using n.c.a. selenium-75 dissolved in
1 mL of benzene and 0.6 mL of DMF containing the copper
catalyst. However, no product was formed under these conditions,
and even increasing both the reaction temperature up to 150 °C
Figure 1. Molecular structure of Ebselen.
and the reaction time up to 14 h did not lead to any formation of
Scheme 1. Radiosynthesis of c.a. [75Se]Ebselen starting from anthranilic acid.19
www.jlcr.org
Copyright © 2015 John Wiley & Sons, Ltd.
J. Label Compd. Radiopharm 2015, 58 141–145

A. Helfer et al.
Scheme 2. Synthesis of Ebselen by ortho-lithiation and subsequent oxidative ring closure.20
[75Se]Ebselen. Thus, the direct transfer of literature conditions21 for
labeling failed again under n.c.a. conditions.
Thus, although the copper-catalyzed synthesis of [75Se]-
Ebselen failed at the n.c.a. level, it was possible with the addition
Therefore, as a further attempt, selenium carrier was added to of selenium carrier which, however, decreases the specific
the reaction mixture. A 0.8 equiv. of selenium relative to the activity considerably. Bhakuni et al.24 reported that the copper-
amount of 2-iodo-N-phenylbenzamide led to the formation of catalyzed reaction is also applicable to the sulfur homologue of
c.a. [75Se]Ebselen with 41% 11% radiochemical yield (RCY) at Ebselen, and even within much shorter reaction time. They
110 °C (Scheme 4). As shown in Figure 2, the RCY of c.a. [75Se]- further reported that an excess of K2CO3 would lower the yield
Ebselen can be increased to 60% 18% at 150 °C, but with very of the corresponding benzoisothiazolone.24 Therefore, the
limited reproducibility.
With regard to the relatively short half-life of selenium-73, the order to obtain [75Se]Ebselen in n.c.a. labeled form.
reaction times of the coupling reaction and the water treatment First experiments using sulfur as non-isotopic carrier under the
addition of sulfur as non-isotopic carrier was also examined in
were reduced to 3 and 1 h, respectively, so that the total reaction same conditions as explored in the c.a. reactions at 110 °C (cf.
time was within one half-life of selenium-73. In contrast to the aforementioned discussion) did not show any product formation.
literature,21 where brine was used to treat the reaction mixture Increasing both the amount of starting material and K2CO3 from
for work up, diluted acetic acid gave more reliable results.
25 to 37.5 μmol, however, led to the formation of the desired
product (Scheme 5).
A study of the influence of the reaction time showed that a
maximum RCY of 55% 7% of n.c.a. [75Se]Ebselen could be
achieved within 3 h, which is far below one half-life of
selenium-73 (Figure 3). The time dependence exhibited a
decrease of the RCY with longer reaction times up to 5 h, most
probably because of decomposition reactions. Final treatment
of the reaction solution with diluted acetic acid was necessary
for about 1 h, which extended the total synthesis time to 4 h.
The sulfur analog exhibits a much higher lipophilicity than
Ebselen. Thus, it was easily possible to develop a reverse phase
HPLC separation as depicted in Figure 4, and n.c.a. [75Se]Ebselen
was obtained in a radiochemical and chemical purity of >99%.
Scheme 3. Proposed mechanism for the N–Se–C cyclization reaction.21
N.c.a. radiosynthesis of Ebselen with the positron emitter
selenium-73
Scheme 4. Copper-catalyzed synthesis of n.c.a. [75Se]Ebselen according to ref. 21
with addition of selenium as isotopic carrier.
When transferring the reaction conditions optimized with
selenium-75 (cf. Scheme 4) to the radiosyntheses with the
positron emitter 73Se, the maximum RCY of n.c.a. [73Se]Ebselen
was limited to approximately 22%. The differences in the RCY
of the same reaction using the two isotopes cannot be explained
by an isotopic effect, but it is presumably caused by changes of
the chemical form of radioselenium in the benzene solution after
separation from the target.
Indeed, a dependence of the labeling yield on the time lapse
between the benzene extraction of radioselenium and the
performance of the labeling step was observed. Thus, an RCY
of about 15% was obtained with labeling reactions immediately
following the extraction of selenium into benzene, but
approximately 50% RCY was obtained with the same benzene
solution used for radiolabeling after 8 days. Comparable yields
of radiolabeling with selenium-75 and selenium-73 were
demonstrated independent of the addition of Se carrier.
Therefore, it can only be concluded that the reactive species in
the benzene solution is formed rather slowly from an unreactive
chemical species, although only one radioactive peak could be
observed in the HPLC analysis of the benzene solution (cf.
Experimental).
Figure 2. Influence of the reaction temperature on the RCY of [75Se]Ebselen
formed by the copper-catalyzed reaction and addition of selenium carrier.
Conditions: 25 μmol of catalyst, 25 μmol of 2-iodo-N-phenylbenzamide, 20 μmol
of selenium, 0.6 mL of DMF, 1 mL of benzene solution of n.c.a. 75Se, 37.5 μmol of
K2CO3, and 3 h reaction time.
J. Label Compd. Radiopharm 2015, 58 141–145
Copyright © 2015 John Wiley & Sons, Ltd.
www.jlcr.org

A. Helfer et al.
Scheme 5. Copper-catalyzed one-pot radiosynthesis of n.c.a. [75Se]Ebselen with sulfur as non-isotopic carrier.
chromatography was carried out with Fluka silica gel 60 (220–440 mesh).
The syntheses of 2-phenyl-1,2-benzisoselenazol-3(2H)-one (Ebselen) and
2-phenyl-1,2-benzisothiozol-3(2H)-one were carried out according to
literature methods.20
Analytical radio-HPLC was performed on a system consisting of a
Hitachi L-6000 pump and a Sykam UV/vis photometer S3300 with a
detector wavelength of 220 nm. Sample injection was accomplished by
a Rheodyne-injector block 7125. For measurement of radioactivity, the
outlet of the UV detector was connected to
a NaI(Tl) well-type
scintillation detector (EG&G ACE Mate Ortec AMETEK GmbH, Germany),
and the recorded data were processed by the software system Raytest
Gina (Raytest, Germany). Radio-TLC was performed on Merck silica gel
plates. The developed TL-chromatograms were measured for
radioactivity on an Instant Imager® (Packard, USA).
Production of n.c.a. [75Se]selenium
Figure 3. Influence of the reaction time on the RCY of [75Se]Ebselen formed by
the copper-catalyzed reaction and addition of sulfur carrier. Conditions: 25 μmol
of catalyst; 37 μmol of 2-iodo-N-phenylbenzamide; 35 μmol of sulfur; 1 mL of
benzene solution of n.c.a. 75Se; 37.5 μmol of K2CO3, 110 °C, and 2 h hydrolysis.
Selenium-75 was produced using 17 MeV protons at the JSW cyclotron
BC-1710 of the Forschungszentrum Jülich via the 75As(p,n)75Se reaction in
a solid Cu3As-target.22 Isolation of the radionuclide was performed as
reported earlier.23 After thermochromatographic separation, n.c.a.
selenium-75 is obtained in oxidized form as [75Se]SeO32À in hydrochloric acid.
It was then reduced with sulfur dioxide to obtain n.c.a. selenium-75, which
was extracted into benzene.22 HPLC analysis showed one uniform peak in
the extraction solution (Luna® 5-μm phenyl–hexyl 100-Å (250 × 10 mm)
column (Phenomenex, Germany); mobile phase consists of MeOH/H2O/
trifluoracetic acid 40:60:0.01 (given as V/V ratios) at a flow rate of 1.0 mL/min).
Production of n.c.a. [73Se]selenium
Selenium-73 was produced through the 75As(p,3n) process on a solid
Cu3As-target using 45 MeV protons at the high energy isochronous
cyclotron (JULIC) of the Forschungszentrum Jülich GmbH.10
Thermochromatographic work up and the following reduction as
described earlier provided an n.c.a. solution of selenium-73 in benzene.
Radiosynthesis of n.c.a. [73,75Se]Ebselen
Figure 4. Separation of Ebselen and its sulfur analog by HPLC. Conditions:
Luna®5 μm phenyl–hexyl 100 Å (150 × 21.2 mm) with MeOH/H2O/trifluoroacetic
acid 40:60:0.1 (v/v/v).
In a 5-mL Wheaton V-Vial, 20 μmol of CuI and 20 μmol of 1,10-
phenanthroline were added in 0.6 mL of dry DMF and stirred for
20 min at room temperature under argon atmosphere. An orange-
brown copper complex was formed that was added to a mixture of
37.5 μmol of 2-iodo-N-phenylbenzamide, 35 μmol of sulfur, and
In the earlier developmental work, the selenium activity
extracted into benzene was considered to be bona fide elemental
radioselenium, because oxidized species were not taken up in the
organic phase after dissolving and reducing the target material. An
37.5 μmol of dry K2CO3 powder in
a 5-mL round-bottom flask.
identification of the definite chemical form has not been carried
Thereafter, the benzene solution with n.c.a. radioselenium was added
to the mixture, the flask equipped with a Dimroth cooler and flushed
with argon. The solution was stirred for 3 h at 110 °C. Afterwards,
22,25
out so far.
Further radioanalytical work on radionuclide
development was out of the scope of this study but appears
mandatory to identify the chemical form of n.c.a. selenium 15 mL of H2O and 2 mL of 1% acetic acid were poured into the
reaction mixture and stirred at room temperature for 1 h. In order
to prevent the loading of HPLC column with particles and copper
salts, an solid phase extraction using SepPak C-18 cartridge was
performed. The cartridge was preconditioned with methanol and
water, and the raw products were eluted with 4 mL of methanol.
Radio-HPLC analysis and separations were performed using a Luna®
5 μm phenyl–hexyl 100 Å (250 × 10 mm) column; mobile phase
present in benzene after extraction from the aqueous solution
of the dissolved target.
Experimental
Material and methods
All chemicals and solvents were purchased from Aldrich (Germany), Fluka consists of MeOH/H2O/trifluoroacetic acid 40:60:0.01 (given as V/V
(Switzerland), and Merck (Germany). They were reagent grade or better ratios) at a flow rate of 1.0 mL/min. The organic solvent of the
and used without further purification. All selenocompounds were product fraction was finally removed by evaporation and neutralized
prepared under
a
slightly positive pressure of argon. Flash
with phosphate-buffered saline to a pH of 7.4.
www.jlcr.org
Copyright © 2015 John Wiley & Sons, Ltd.
J. Label Compd. Radiopharm 2015, 58 141–145

A. Helfer et al.
[5] T. Blum, J. Ermert, W. Wutz, D. Bier, H. H. Coenen, J Label Compd
Radiopharm 2004, 47, 415.
[6] R. Riemsma, M. Al, I. Corro Ramos, S. Deshpande, N. Armstrong, Y. C.
Lee, S. Ryder, C. Noake, M. Krol, M. Oppe, J. Kleijnen, H. Severens,
Health Technol Assess 2013, 17, 1.
[7] M. Neves, A. Kling, A. Oliveira, J Radioanal Nucl Chem 2005, 266, 377.
[8] A. Graebert, D. Schmidt, A. Kyriakopoulos, Trace Elem Electrolytes
2010, 27, 140.
Conclusions
In an initial study, the preparation of [75Se]Ebselen by a copper-
catalyzed one-pot radiosynthesis could only be achieved under
c.a. conditions but in RCYs of 60% 18%. The n.c.a. product
could then be obtained using sulfur as non-isotopic carrier in
the reaction mixture and its separation from its co-produced
sulfur analog by HPLC. After optimization of reaction parameters,
n.c.a. [75Se]Ebselen could be synthesized at 110 °C with RCYs of
55% 7% within 3 h. On transferring the conditions to the
[9] S. M. Qaim, Radiochim Acta 2011, 99, 611.
[10] A. Mushtaq, S. M. Qaim, G. Stöcklin, Int J Appl Radiat Isot 1988,
39, 1085.
[11] T. Blum, J. Ermert, H. H. Coenen, Appl Radiat Isot 2002, 57, 51.
radiosyntheses with the positron emitter 73Se, however, n.c.a. [12] T. Blum, J. Ermert, H. H. Coenen, Nucl Med Biol 2003, 30, 361.
[73Se]Ebselen could only be obtained with an RCY of 22% 1%.
Nevertheless, the compound can now be used as a potential
radiotracer in preclinical evaluation studies aiming at tracer
application with PET.
[13] M. Parnham, H. Sies, Expert Opin Investig Drugs 2000, 9, 607.
[14] M. J. Parnham, H. Sies, Biochem Pharmacol 2013, 86, 1248.
[15] T. Yamaguchi, K. Sano, K. Takakura, I. Saito, Y. Shinohara, T. Asano, H.
Yasuhara, Stroke 1998, 29, 12.
[16] H. Kaur, A. Prakash, B. Medhi, Pharmacology 2013, 92, 324.
[17] S. Park, S. Kang, D. S. Kim, B. K. Shin, N. R. Moon, J. W. Daily Iii, Free
Radic Res 2014, 48, 864.
[18] N. Singh, A. C. Halliday, J. M. Thomas, O. V. Kuznetsova, R. Baldwin, E.
C. Y. Woon, P. K. Aley, I. Antoniadou, T. Sharp, S. R. Vasudevan, G. C.
Churchill, Nat Commun 2013, 4, 1332.
Conflict of Interest
The authors did not report any conflict of interest.
[19] R. Cantineau, G. Tihange, A. Plenevaux, J Label Compd Radiopharm
1986, 23, 59.
References
[20] L. Engman, A. Hallberg, J Org Chem 1989, 54, 2964.
[21] S.J.Balkrishna,B.S.Bhakuni,D.Chopra,S.Kumar,OrgLett2010,12,5394.
[22] G. Blessing, N. Lavi, S. M. Qaim, Int J Appl Radiat Isot 1992, 43, 455.
[23] G.Blessing,N.Lavi,K.Hashimoto,S.M.Qaim,RadiochimActa1994,65,93.
[24] B. S. Bhakuni, S. J. Balkrishna, A. Kumar, S. Kumar, Tetrahedron Lett
2012, 53, 1354.
[1] K. Ekambaramgnanadesigan, E. Balumahendran, N. Gnanagurudasan,
S. Kumar, Int J Pharma Bio Sci 2013, 4, B1.
[2] C. Galinha, M. C. Freitas, A. M. G. Pacheco, J. Coutinho, B. Maçãs, A. S.
Almeida, J Radioanal Nucl Chem 2012, 291, 193.
[3] T. W. Gebel, Int J Hyg Environ Health 2002, 205, 505.
[4] J. Ermert, T. Blum, K. Hamacher, H. H. Coenen, Radiochim Acta 2001,
89, 863.
[25] A. Plenevaux, M. Guillaume, C. Brihaye, C. Lemaire, R. Cantineau, Appl
Radiat Isot 1990, 41, 829.
J. Label Compd. Radiopharm 2015, 58 141–145
Copyright © 2015 John Wiley & Sons, Ltd.
www.jlcr.org